(Supported by the NIH/NCRR/P41 RR 01219 grant). Automated (or semi-automated) electron tomography is required to record tomographic tilt series with minimal specimen irradiation. This is essential for frozen-hydrated specimens, and also prevents specimen distortion of plastic-embedded specimens during the tilt series. The Tietz (TVIPS, Gauting Germany) slow-scan CCD system and automated tomography software were installed on the IVEM, and the software was adapted to our microscope (Rath, B.K., Marko, M., Radermacher, M., Frank, J. (1997) Low-dose automated electron tomography a recent implementation. J. Str. Biol. 120, 210-218). After much experience with cryo-tomography of the mitochondria and axoneme (see above TRD projects), recording of cryo tilt series is now nearly routine. A new calibration procedure to correct beam shift and change in illumination size due to focus change was added to the Tietz software. This can be applied to both semi-automated and automated tomography. This was required because of the poor performance of the goniometer with respect to focus change (see TRD subproject project "IVEM maintenance and development"). In most of our tomography applications, we find that limits on the size of an object that can be reconstructed at a given resolution need to be increased. Since we will not be able to afford a CCD camera with a larger array until the next renewal of this grant, we are investigating low-cost alternatives to acquire larger images without resorting to the use of photographic film. We obtained automatic image montaging or "tiling" software from the Tietz company, but are having difficulties in implementing it on our system, which is of an earlier generation than the one it was written for. Hardware is in place for addition of STEM imaging to tomography, using our existing STEM unit. This will give us images up to 2048x2048 pixels in size. We purchased a Pentek A/D converter board for the Tietz computer, and the connections to the STEM unit (video signal, vertical blanking pulse, and scan initiation switch) are made. We are waiting for information from the Tietz company, needed for i ntegration of the Pentek board with the Tietz software. Improvements were made in the single- and double-tilt alignment software, reducing alignment errors. Html documentation on the operation of the IVEM for low-dose imaging and tomography work was written for the use of potential new users. This can be found at: http://www.wadsworth.org/spider_doc/spider/docs/techs/lgstr/index.html